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. 1984 Jul 1;221(1):113–120. doi: 10.1042/bj2210113

Rapid optimization of immunoadsorbent characteristics.

J Janatova, R J Gobel
PMCID: PMC1144009  PMID: 6380495

Abstract

Immunoaffinity chromatography is employed in many research areas. We have developed an assay system that overcomes some of the tediousness and uncertainty in dealing with immunoadsorbents (IA). The preparation of IA and the effect of various procedures on the dissociation of antigen-antibody complex and the regeneration of IA are rapidly screened by polyacrylamide-gel electrophoresis in the presence of sodium dodecyl sulphate. Non-covalently bound proteins are dissociated and separated during the electrophoresis from IA beads. A wide variety of associative and dissociative conditions can be tested on small amounts of IA. Information about biospecifially and non-biospecifically adsorbed proteins can also be obtained. By using relatively small volumes of media containing either an antigen or another biospecific molecule, the optimal parameters for affinity chromatography (specificity, binding capacity, efficiency of solvents in dissociation of the complex and their effect on the adsorbent), or even for ion-exchange chromatography, can be determined without first performing several time- and material-consuming chromatographic experiments.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Campbell D. H., Luescher E., Lerman L. S. Immunologic Adsorbents: I. Isolation of Antibody by Means of a Cellulose-Protein Antigen. Proc Natl Acad Sci U S A. 1951 Sep;37(9):575–578. doi: 10.1073/pnas.37.9.575. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Cheson B. D., Walker H. S., Heath M. E., Gobel R. J., Janatova J. Defective binding of the third component of complement (C3) to Streptococcus pneumoniae in multiple myeloma. Blood. 1984 Apr;63(4):949–957. [PubMed] [Google Scholar]
  3. Cuatrecasas P. Affinity chromatography of macromolecules. Adv Enzymol Relat Areas Mol Biol. 1972;36:29–89. doi: 10.1002/9780470122815.ch2. [DOI] [PubMed] [Google Scholar]
  4. Cuatrecasas P. Protein purification by affinity chromatography. Derivatizations of agarose and polyacrylamide beads. J Biol Chem. 1970 Jun;245(12):3059–3065. [PubMed] [Google Scholar]
  5. Dean P. D., Brown P., Leyland M. J., Watson D. H., Angal S., Harvey M. J. Electrophoretic desorption of affinity adsorbents [proceedings]. Biochem Soc Trans. 1977;5(4):1111–1113. doi: 10.1042/bst0051111a. [DOI] [PubMed] [Google Scholar]
  6. GURVICH A. E., DRIZLIKH G. I. USE OF ANTIBODIES ON AN INSOLUBLE SUPPORT FOR SPECIFIC DETECTION OF RADIOACTIVE ANTIGENS. Nature. 1964 Aug 8;203:648–649. doi: 10.1038/203648a0. [DOI] [PubMed] [Google Scholar]
  7. Janatova J., Tack B. F. Fourth component of human complement: studies of an amine-sensitive site comprised of a thiol component. Biochemistry. 1981 Apr 28;20(9):2394–2402. doi: 10.1021/bi00512a005. [DOI] [PubMed] [Google Scholar]
  8. Janatova J. The third (C3) and the fourth (C4) components of complement: labile binding site and covalent bond formation. Ann N Y Acad Sci. 1983;421:218–234. doi: 10.1111/j.1749-6632.1983.tb18111.x. [DOI] [PubMed] [Google Scholar]
  9. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  10. MacSween J. M., Eastwood S. L. Recovery of antigen from staphylococcal protein A--antibody adsorbents. Methods Enzymol. 1981;73(Pt B):459–471. [PubMed] [Google Scholar]
  11. O'carra P., Barry S., Griffin T. Interfering and complicating adsorption effects in bioaffinity chromatography. Methods Enzymol. 1974;34:108–126. doi: 10.1016/s0076-6879(74)34011-6. [DOI] [PubMed] [Google Scholar]
  12. Parikh I., March S., Cuatercasas P. Topics in the methodology of substitution reactions with agarose. Methods Enzymol. 1974;34:77–102. doi: 10.1016/s0076-6879(74)34009-8. [DOI] [PubMed] [Google Scholar]
  13. Reid K. B., Porter R. R. The proteolytic activation systems of complement. Annu Rev Biochem. 1981;50:433–464. doi: 10.1146/annurev.bi.50.070181.002245. [DOI] [PubMed] [Google Scholar]
  14. Robbins J. B., Schneerson R. Immunoadsorbents. Methods Enzymol. 1974;34:703–722. doi: 10.1016/s0076-6879(74)34093-1. [DOI] [PubMed] [Google Scholar]
  15. Silman I., Katchalski E. Water-insoluble derivatives of enzymes, antigens, and antibodies. Annu Rev Biochem. 1966;35:873–908. doi: 10.1146/annurev.bi.35.070166.004301. [DOI] [PubMed] [Google Scholar]
  16. Wilchek M., Jakoby W. B. The literature on affinity chromatography. Methods Enzymol. 1974;34:3–10. doi: 10.1016/s0076-6879(74)34004-9. [DOI] [PubMed] [Google Scholar]
  17. Zoller M., Matzku S. Antigen and antibody purification by immunoadsorption: elimination of non-biospecifically bound proteins. J Immunol Methods. 1976;11(3-4):287–295. doi: 10.1016/0022-1759(76)90122-8. [DOI] [PubMed] [Google Scholar]

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